Adeno-associated virus (AAV) is a non-pathogenic human parvovirus, and recombinant AAV vectors have gained significant attention owing to their safety, and in some cases, clinical efficacy, in a number of Phase I/II clinical trials. However, relatively large vector doses are needed to achieve clinical efficacy, and such high doses can lead to host immune responses. The main goal of the proposed research is to develop recombinant AAV vectors that are not only highly efficient, but are also effective at lower doses, thereby reducing the probability of inducing the host cell immune responses, the vector production costs, as well as the economic cost per patient, for the potential gene therapy of a wide variety of human diseases. Dr. Arun Srivastava (PI, University of Florida) has developed the next generation (`NextGen') of AAV vectors by modifying the AAV capsid protein, and these vectors work at significantly reduced doses, and are less immunogenic. His lab has also modified the AAV vector genome to develop the generation X (`GenX') AAV vectors that further enhance the transgene expression. As a result, Dr. Srivastava has partnered with Medosome Biotec, LLC (a Florida-based small business focused on the development and commercialization of technologies that enrich the lives of children by providing solutions to childhood diseases, disabilities and deficiencies) to further develop and commercialize this vector. The team hypothesizes that encapsidation of the GenX AAV genomes into NextGen AAV capsids will lead to the generation optimized (Opt) AAV vectors, with which significantly higher transduction efficiency at further reduced doses will be achieved. The safety and efficacy of these novel OptAAV serotype vectors will be evaluated in xenograft and humanized mice models prior to their potential use in human gene therapy. In the proposed Phase I STTR application, the following two specific aims will be pursued: Specific Aim 1: Development of Opt AAV serotype vectors in which GenX AAV genomes are encapsidated in NextGen capsids Specific Aim 2: Evaluation of safety and efficacy of the Opt AAV serotype vectors in xenograft and humanized murine models in vivo The successful completion of these studies will not only yield new insights into basic molecular biology of AAV, but will also be directly applicable in the further development of a vast repertoire of novel recombinant AAV serotype vectors for their optimal use in human gene therapy.